The aim of this proposal is to elucidate T cell receptor (TCR) crossreactivity at the molecular level. The unique model developed involves crossreactive TCR recognition of the immunodominant viral peptide coupled to two very closely related class I MHC variants. TCRs to be studied share the same TCRbeta chains, but some see only one of the complexes (monospecific recognition) while others see both (crossreactive recognition). This indicates that the TCRalpha chain must play a decisive role in crossreactivity. TCR sequencing, generation and analysis of single TCR chain transgenic (scTCRTg mice), Crystal structure determination and TCR mutagenesis will be used to dissect the molecular contacts that permit crossreactivity, a phenomenon underlying alloreactivity, tolerance induction, transplant rejection and autoimmunity. The aims are: 1. To analyze functional reactivity and TCR sequence of crossreactive and monospecific CTL clones from normal and scTCRTg mice and establish the preliminary contacts that define crossreactive and monospecific TCR:peptide:MHC interactions for closely related TCRs. 2. To determine the x-ray structures of the monospecific and crossreactive TCRs with the appropriate pep:MHC complexes, and use site-directed mutagenesis to definitively dissect crossreactive recognition.